Project summary: Immune recovery after bone marrowtransplantation remains a significant clinical problem, one that may be addressed by progenitor cell transplants aimed at augmenting lymphoid recovery. Recent studies using clonal cultures and intrathymic transplant models have defined the point at which lymphoid progenitors lose myeloid potential, as well as the stage at which the B lineage becomes specified and irreversible. However, the clonal approaches utilized to date for defining lymphoid specification suffer from the criticism that they lack physiological significance. As a result, the primary physiological pathways for T and B lineage development remain incompletely characterized. We have defined a series of lymphoid progenitors in mouse bone marrow based on expression of L-selectin and other antigens. L-selectin-neg cells are more robust in engraftment experiments, while L-selectin-pos cells are more restricted to the T lineage. In the next funding period we seek to define the lineage relationship and developmental potential of these two populations of adult bone marrow-derived progenitors in comparison to previously defined lymphoid progenitor populations. We also seek to resolve conflicting data regarding lymphoid specification by making direct comparisons between transplant and culture model systems. We aim to test the hypothesis that the most relevant proximal progenitor for the T and B lineages is a multipotent progenitor population in bone marrow. We seek to define and isolate these cells, as well as their direct progeny that are more restricted in their developmental potential. We will utilize clonal viral marking of prospectively isolated progenitor populations in order to track efficiency and clonality of reconstitution following intravenous transplantation into normal as well as irradiated mice. By combining our expertise in stem cell biology with our interest in early lymphoid development, we aim to achieve a thorough understanding of where lymphoid and myeloid pathways diverge in physiologically relevant models that may find applications in clinical bone marrowtransplantation. Relevance: The immune response is critical to maintaining a normal lifestyle. In certain disease states, or following treatment for cancer, the immune system is weakened. This work seeks to define transplantation interventions that will reduce the complications associated with immune deficiencies.